The proposed research is aimed at understanding the mechanism of heme transport from one hepatic compartment to another and the factors which govern the intracellular heme distribution among the different heme-containing compartments such as cytosol, microsomes and mitochondria. Using a dual radioisotope pulse method employing the 3H- and 14C-labelled heme precursor delta-aminolevulinic acid (ALA), we will initially identify the cytosolic proteins of rat liver which are implicated as carriers in the dynamic process of heme transport. Those cytosolic proteins which function in the intracellular transport of heme and/or its precursors, will be purified and monospecific antibodies will be developed to them. With an immunoprecipitation method the results obtained by conventional protein separation techniques will be verified and further quantitated. Other lines of investigation will define the common and separate pathways for the intracellular transport, distribution and utilization of heme produced in the liver as well as that intravenously administered. Once the molecular events of heme distribution in untreated animals are better understood, we will assess the influence of altered iron and porphyrin metabolism on intracellular heme transport. Hepatic heme levels will be altered by inhibiting heme synthesis through limiting iron supply or by blocking enzymatic steps of its synthesis, e.g. with porphyrogen administration. The heme levels will also be altered by increasing the cellular heme demand by induction of microsomal cytochrome P-450 or by decreasing microsomal heme levels by accelerating the catabolism of cytochrome P-450. The proposed work has as its ultimate goal to furnish the necessary information on the physiology of intracellular heme distribution and transport and the consequences that aberrations of iron and porphyrin metabolism have on this process. This information is a prerequisite for the rationale of heme infusions in treatment of porphyrias.